Do corals form their skeletons by precipitation from solution or by attachment of amorphous precursor particles as observed in other minerals and biominerals? The classical model assumes precipitation in contrast with observed “vital effects,” that is, deviations from elemental and isotopic compositions at thermodynamic equilibrium. Here, we show direct spectromicroscopy evidence in Stylophora pistillata corals that two amorphous precursors exist, one hydrated and one anhydrous amorphous calcium carbonate (ACC); that these are formed in the tissue as 400-nm particles; and that they attach to the surface of coral skeletons, remain amorphous for hours, and finally, crystallize into aragonite (CaCO3). We show in both coral and synthetic aragonite spherulites that crystal growth by attachment of ACC particles is more than 100 times faster than ion-by-ion growth from solution. Fast growth provides a distinct physiological advantage to corals in the rigors of the reef, a crowded and fiercely competitive ecosystem. Corals are affected by warming-induced bleaching and postmortem dissolution, but the finding here that ACC particles are formed inside tissue may make coral skeleton formation less susceptible to ocean acidification than previously assumed. If this is how other corals form their skeletons, perhaps this is how a few corals survived past CO2 increases, such as the Paleocene–Eocene Thermal Maximum that occurred 56 Mya.

Aragonite skeletons in corals are key contributors to the storage of atmospheric CO2 worldwide. Hence, understanding coral biomineralization/calcification processes is crucial for evaluating and predicting the effect of environmental factors on this process. While coral biomineralization studies have focused on adult corals, the exact stage at which corals initiate mineralization remains enigmatic. Here, we show that minerals are first precipitated as amorphous calcium carbonate and small aragonite crystallites, in the pre-settled larva, which then evolve into the more mature aragonitic fibers characteristic of the stony coral skeleton. The process is accompanied by modulation of proteins and ions within these minerals. These findings may indicate an underlying bimodal regulation tactic adopted by the animal, with important ramification to its resilience or vulnerability toward a changing environment.

In reef-building corals, larval settlement and its rapid calcification provides a unique opportunity to study the bio-calcium carbonate formation mechanism involving skeleton morphological changes. Here we investigate the mineral formation of primary polyps, just after settlement, in two taxa of the pocilloporoid corals: Stylophora pistillata (Esper, 1797) and Pocillopora acuta (Lamarck, 1816). We show that the initial mineral phase is nascent Mg-Calcite, with rod-like morphology in P. acuta, and dumbbell morphology in S. pistillata. These structures constitute the first layer of the basal plate which is comparable to Rapid Accretion Deposits (Centers of Calcification, CoC) in adult coral skeleton. We found also that the rod-like/dumbbell Mg-Calcite structures in subsequent growth step may merge into larger aggregates by deposition of aragonite needles. Our results suggest that a biologically controlled mineralization of initial skeletal deposits (CoC) occurs in three steps: first, vesicles filled with divalent ions are formed intracellularly. These vesicles are then transferred to the calcification site, forming nascent Mg-Calcite rod/pristine dumbbell structures. During the third step, aragonite crystals may develop between these structures forming spherulite-like aggregates.

Depth related parameters, specifically light, affect different aspects of corals physiology, including fluorescence. Green fluorescence protein (GFP)-like pigments found in many coral species have been suggested to serve a variety of functions, including photo-protection and photo-enhancement. Using fluorescence imaging and molecular analysis, we further investigated the role of these proteins on the physiology of the coral Stylophora pistillata and its algal partners. Fluorescence was found to differ significantly between depths for larvae and adult colonies. Larvae from the shallow reef presented a higher GFP expression and a greater fluorescence intensity compared to the larvae from the mesophotic reef, reflecting the elevated need for photo-protection against high light levels characteristic of the shallow reef, thus supporting the "sunscreen" hypothesis. Additionally, given the lower but still occurring protein expression under non-damaging low light conditions, our results suggest that GFP-like proteins might act to regulate the amount of photosynthetically usable light for the benefit of the symbiotic algae. Moreover, we propose that the differences in GFP expression and green fluorescence between shallow and deep larvae indicate that the GFPs within coral larvae might serve to attract and retain different symbiont clades, increasing the chances of survival when encountering new environments.

In light of the chronic stress and mass mortality reef-building corals face under climate change, it is critical to understand the processes essential to reef persistence and replenishment, including coral reproduction and development. Here we quantify gene expression and size sensitivity to ocean acidification across a set of developmental stages in the rice coral, Montipora capitata. Gametes and then embryos and swimming larvae were exposed to three pH treatments ranging from 7.8 (Ambient), 7.6 (Low) and 7.3 (Xlow) from fertilization to 9 days post-fertilization. Embryo development and size, planula volume, and stage-specific gene expression were compared between treatments at each stage to determine the effects of acidified seawater on early development. While there was no measurable size differentiation between fertilized eggs and embryos at the prawn chip stage exposed to ambient, low, and extreme low pH, early gastrula and planula raised in reduced pH treatments were significantly smaller than those raised in ambient seawater, suggesting an energetic cost to developing under low pH. However, no differentially expressed genes emerged between treatments at any time point, except swimming larvae. Larvae from pH 7.6 showed upregulation of genes involved in cell division, regulation of transcription, lipid metabolism, and oxidative stress in comparison to the other two treatments, and smallest sizes in this treatment. While low pH appears to increase energetic demands and trigger oxidative stress, the developmental process is robust to this at a molecular level, with swimming larval stage reached in all pH treatments.

No abstract

In the tropical and subtropical oceans, corals provide a literal and figurative ecological framework that retains nutrients, supports high rates of primary production, and permits extensive biological diversity. These fragile ecosystems are threatened with extinction in the coming century, in part due to the acidification of the ocean [1][2]. The change in the acidity of the ocean may affect the coral skeleton formation, which is composed of calcium carbonate mineral. Elucidating the mechanism of calcium carbonate mineral formation in corals may help us to understand how environmental changes affect the process of coral biomineralization. Although various aspects of biomineralization in corals have been studied for decades, mostly on adult corals, the basic mechanism responsible for the precipitation of calcium carbonate in the form of aragonite remains enigmatic [reviewed by 3].Corals have a biphasic life cycle with planktonic larval stages and benthic adults. These two phases are separated by settlement and metamorphosis, which are critical stages in coral development during which planulae undergo intense changes in morphology, building of new tissues, initiation of calcium carbonate precipitation, and in some cases uptake of symbionts. The planktonic free-swimming planulae deposit minerals almost immediately after settlement [4]. This suggests that immature mineral phases (presumably amorphous calcium carbonate -ACC) is present in pre-settled planulae.To elucidate the key mechanism that facilitates the initial, rapid calcification in the early stages of the coral development, we correlate cryo-scanning electron microscopy (SEM) with cryo-energy-dispersive X-ray spectroscopy (EDS) and cryo-fluorescence techniques on coral planulae frozen samples [5]. Using this approach and thus, avoiding the process of chemical fixation, dehydration and chemical staining with heavy metals, each planula is rapidly frozen at high pressure. As a result, the water is vitrified while keeping the sample as close as possible to its native state. We observed freeze-fractured surfaces of the sample using the cryo-SEM with both secondary electron and backscattered electron (BSE) detectors. The elements that are present in the mineralized regions are detected by EDS under cryogenic conditions. The cryo-fluorescence platform helps to identify auto-fluorescent symbionts and to detect mineralized regions in the developing planula via calcein blue staining.Our results show that first mineral deposition in corals already starts at the pre-settlement (Fig. 1). Immature minerals can vary in shape, Mg content and crystallinity. After settling, the aboral epidermis is attached to the substrate and begins skeleton formation. The first calcareous elements after settling are circular platelets and rod-shaped granules, which can also vary in Mg and Ca content and aggregate to form the primary septa, followed by the formation of the basal disk [6,7].

Scleractinian corals are evolutionary-successful calcifying marine organisms, which utilize an endo-symbiotic relationship with photosynthetic dinoflagellate algae that supply energy products to their coral hosts. This energy further supports a higher calcification rate during the day in a process known as light enhanced calcification. Although this process has been studied for decades, the mechanisms behind it are still unknown. However, photosynthesis and respiration also cause daily fluctuations in oxygen and pH levels, resulting in the coral facing highly variable conditions. Here we correlated gene expression patterns with the physiological differences along the diel cycle to provide new insights on the daily dynamic processes, including circadian rhythm, calcification, symbiosis, cellular arrangement, metabolism, and energy budget. During daytime, when solar radiation levels are highest, we observed increased calcification rate combined with an extensive up-regulation of genes associated with reactive oxygen species, redox, metabolism, ion transporters, skeletal organic matrix, and mineral formation. During the night, we observed a vast shift toward up-regulation of genes associated with cilia movement, tissue development, cellular movement, antioxidants, protein synthesis, and skeletal organic matrix formation. Our results suggest that light enhanced calcification is related to several processes that occur across the diel cycle; during nighttime, tissue might elevate away from the skeleton, extending the calcifying space area to enable the formation of a new organic framework template. During daytime, the combination of synthesis of acid-rich proteins and a greater flux of ions to the sites of calcification facilitate the conditions for extensive mineral growth.